"Functional Analysis of Kox 1: A Human Zinc Finger Gene" proposes a molecular characterization of the structure and function of Kox 1. Kox 1 is one of 30 unique genes isolated from human leukemia cDNA libraries (1), which contain regions of amino acid sequence homology with Kruppel type zinc finger genes of other species (3,4,5). Kruppel type zinc finger genes often have transcriptional regulatory abilities and are important in the control of cell growth and differentiation (3,5). We have recently described a transcriptional repression domain in Kox 1 (4). This is the only known function of a Kox gene. Kox 1 was chosen for this analysis because: 1) it is expressed primarily in hematopoietic cells, 2) a full length cDNA is available; and 3) the domain we described in the amino (NH2) portion of the protein which mediates transcriptional repression contains a KRAB (KRuppel Associated Box) which may represent a protein/protein interactive domain. There is a strong correlation between oncogenes, tumor suppressor genes and transcription factors. This correlation along with the general role of the Kruppel genes in development suggest that this project will contribute to the understanding of both normal hematopoietic/lineage development, and leukemogenesis. The Specific Aims are: 1) To define the minimal protein domain that is both necessary and sufficient for the transcription repression activity. This will be done using the deletion strategy already successfully used in our preliminary Kox 1 analysis; 2) To identify proteins(s) which may interact with Kox 1, with both differential immunoprecipitation and chromatography techniques; 3) To identify the specific DNA binding site(s) recognized by the eleven zinc fingers of Kox 1. This will be done using a library of synthetic degenerative oligonucleotides, PCR, and gel retardation experiments; 4) To identify target gene(s) regulated by Kox 1. Once binding sites are identified, we will both probe genomic DNA libraries and search the existing sequence databanks for potential target genes. 5) To examine human leukemia and lymphoma specimens for evidence of expression and mutation of Kox 1. A combination of Southern and Northern blots, and SSCP techniques will be used.